Abstract: The pathogenesis of many diseases is closely related to enzymes, and the development of enzyme inhibitors is also increasing exponentially. The electrochemical biosensor constructed with composite enzyme electrode can analyze and detect enzyme inhibitor drugs effectively and efficiently. According to the various characteristics of the electrode materials, single or composite materials are selected for immobilizing the enzymes, and then a biorecognition receptor is formed to detect the enzyme inhibition activity of the analyte. In this review, we summarized the advances of the biosensors in the detection of enzyme inhibitor drugs in the past five years in terms of target enzymes that are closely related to human diseases and drug mechanisms, which provides a theoretical basis for further screening of small molecule lead compounds as enzyme inhibitors.